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First molecular and biochemical analysis of in vivo affinity maturation in an ectothermic vertebrate.

ABSTRACT: The cartilaginous fish are the oldest phylogenetic group in which Igs have been found. Sharks produce a unique Ig isotype, IgNAR, a heavy-chain homodimer that does not associate with light chains. Instead, the variable (V) regions of IgNAR bind antigen as soluble single domains. Our group has shown that IgNAR plays an integral part in the humoral response of nurse sharks (Ginglymostoma cirratum) upon antigen challenge. Here, we generated phage-displayed libraries of IgNAR V regions from an immunized animal and found a family of clones derived from the same rearrangement event but differentially mutated during expansion. Because of the cluster organization of shark Ig genes and the paucicopy nature of IgNAR, we were able to construct the putative ancestor of this family. By studying mutations in the context of clone affinities, we found evidence that affinity maturation occurs for this isotype. Subsequently, we were able to identify mutations important in the affinity improvement of this family. Because the family clones were all obtained after immunization, they provide insight into the in vivo maturation mechanisms, in general, and for single-domain antibody fragments.

SUBMITTER: Dooley H

PROVIDER: S-EPMC1413636 | biostudies-literature | 2006 Feb

REPOSITORIES: biostudies-literature

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First molecular and biochemical analysis of in vivo affinity maturation in an ectothermic vertebrate.

Dooley Helen H Stanfield Robyn L RL Brady Rebecca A RA Flajnik Martin F MF

Proceedings of the National Academy of Sciences of the United States of America 20060130 6

The cartilaginous fish are the oldest phylogenetic group in which Igs have been found. Sharks produce a unique Ig isotype, IgNAR, a heavy-chain homodimer that does not associate with light chains. Instead, the variable (V) regions of IgNAR bind antigen as soluble single domains. Our group has shown that IgNAR plays an integral part in the humoral response of nurse sharks (Ginglymostoma cirratum) upon antigen challenge. Here, we generated phage-displayed libraries of IgNAR V regions from an immun ...[more]

PMID: 16446445

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Project description:Transcriptional profiling of gene responses in liver in the coral reef fish Pomacentrus moluccensis in response to different types of environmental stress: cold, heat, hypoxic and hyposmotic shock. Goal was to determine the common effects of different types of environmental stress on gene expression as well as responses unique to different stressors. Abstract from Kassahn et al. BMC Genomics (2007) 8:358 Background While our understanding of the importance of transcriptional regulation for biological function is continuously growing, we still know comparatively little about how environmentally-induced stress affects gene expression in vertebrates and how consistent transcriptional stress responses are across different types of environmental stress. Results In this study, we looked for a genetic measure of environmental stress and used a multi-stressor approach to identify components of a common stress response as well as components unique to different types of environmental stress. We exposed individuals of the coral reef fish Pomacentrus moluccensis to hypoxic, hyposmotic, cold and heat shock and measured the responses of approximately 16,000 genes in liver. We also compared winter and summer responses to heat shock to examine the capacity for such responses to vary with acclimation to different ambient temperatures. We identified a series of gene functions that were consistently involved in all stress responses examined here, suggesting common effects of stress on biological function. These common responses were achieved by the regulation of largely independent sets of genes and the responses of individual genes varied greatly across different stress types. However, we were able to identify groups of co-regulated genes, the genes within which shared similar functions. Given current estimates of climatic change, we were particularly interested in the response to prolonged heat exposure. In total, 324 gene loci were differentially expressed following exposure to heat over five days. The functions of these heat-responsive genes suggest that prolonged heat stress leads to oxidative stress and protein damage, challenge of the immune system, and a re-allocation of energy sources. Conclusion This is the first environmental genomic study to measure gene regulation in response to different environmental stressors in a natural population of a warm-adapted ectothermic vertebrate. This study offers insight into the effects of environmental stress on biological function and sheds light on the expected sensitivity of coral reef fishes to elevated temperatures in the future. Keywords: Stress response

Dataset Information

First molecular and biochemical analysis of in vivo affinity maturation in an ectothermic vertebrate.

Publications

First molecular and biochemical analysis of in vivo affinity maturation in an ectothermic vertebrate.

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